This is a proposal to examine the role of Ca in regulating adenylyl cyclase(AC) activity in chick cardiac myocytes. Evidence is presented that the myocytes express type V nd VI AC isoforms, both of which are inhibited by Ca. The PI has found that lowering external Ca stimulates cAMP production in response to either ISO or forskolin. The applicant proposes to identify the AC isoforms at the protein level using isoform- specific antibodies, and to alter expression levels of types V and VI isozymes in embryonic chick heart myocytes using adenovirus-mediated expression techniques (sense or antisense), an to examine the effects of these alterations on the Ca-AC interaction. AC activities will be measured in myocytes expressing known levels of AC V and VI, and the effects of Ca will be measured in intact cells (variations in extracellular Ca+/- nifedipine), in permeabilized cells and in isolated membranes. The second specific aim will focus on the long term effects of exposure to Ca channel blockers on myocyte AC levels, using a CRE- luciferase reporter construct, to assess cellular cAMP levels. The third specific aim will investigate potential phosphorylation of th AC isoforms by PKA and PKC, and associated effects on activity levels and the"cross- talk" between Ca and cAMP production.